Biomedical subjects

C Sternini

Publications and source records attributed to C Sternini.

At least 37 records · Page 2Linked to original sources

PubMed1999

Expression of cholecystokinin A receptors in neurons innervating the rat stomach and intestine.

BACKGROUND & AIMS: Two distinct receptors, cholecystokinin (CCK)-A and CCK-B, mediate CCK effects in the digestive system. The aim of this study was to elucidate the cellular sites of expression of CCK-A receptor in the rat stomach and small intestine. METHODS: We developed and characterized an antibody to the N-terminal region (LDQPQPSKEWQSA) of rat CCK-A receptor and used it for localization studies with immunohistochemistry. RESULTS: Specificity of the antiserum was demonstrated by (1) detection of a broad band at 85-95 kilodaltons in Western blots of membranes from CCK-A receptor CHO-transfected cells; (2) cell surface staining of CCK-A receptor-transfected cells, (3) translocation of CCK-A receptor immunostaining in CCK-A receptor-transfected cells after exposure to CCK; and (4) abolition of tissue immunostaining by preadsorbtion of the antibody with the peptide used for immunization. CCK-A receptor immunoreactivity was localized to myenteric neurons and to fibers in the muscle and mucosa. In the stomach, myenteric neurons and mucosal fibers were abundant. Many CCK-A receptor myenteric neurons contained the inhibitory transmitter vasoactive intestinal polypeptide, and some were immunoreactive for the excitatory transmitter substance P. Subdiaphragmatic vagotomy reduced the density of CCK-A receptor fibers in the gastric mucosa by approximately 50%, whereas celiac/superior mesenteric ganglionectomy had no detectable effect on fiber density. CONCLUSIONS: CCK-A receptor is expressed in functionally distinct neurons of the gastrointestinal tract. CCK-A receptor may mediate reflexes stimulated by CCK through the release of other transmitters from neurons bearing the receptor.

Amino Acid Sequence↗

PubMed1999

Activation and internalization of the mu-opioid receptor by the newly discovered endogenous agonists, endomorphin-1 and endomorphin-2.

The multiple effects of opiate alkaloids, important therapeutic drugs used for pain control, are mediated by the neuronal miro-opioid receptor. Among the side effects of these drugs is a profound impairment of gastrointestinal transit. Endomorphins are opioid peptides recently isolated from the nervous system, which have high affinity and selectivity for micro-opioid receptors. Since the miro-opioid receptor undergoes ligand-induced receptor endocytosis in an agonist-dependent manner, we compared the ability of endomorphin-1, endomorphin-2 and the micro-opioid receptor peptide agonist, [D-Ala2,MePhe4,Gly-ol5]-enkephalin (DAMGO), to induce receptor endocytosis in cells transfected with epitope-tagged micro-opioid receptor complementary DNA, and in myenteric neurons of the guinea-pig ileum, which naturally express this receptor. Immunohistochemistry with antibodies to the FLAG epitope or to the native receptor showed that the micro-opioid receptor was mainly located at the plasma membrane of unstimulated cells. Endomorphins and DAMGO induced micro-opioid receptor endocytosis into early endosomes, a process that was inhibited by naloxone. Quantification of surface receptors by flow cytometry indicated that endomorphins' and DAMGO stimulated endocytosis with similar time-course and potency. They inhibited with similar potency electrically induced cholinergic contractions in the longitudinal muscle-myenteric plexus preparation through an action antagonized by naloxone. The apparent affinity estimate of naloxone (pA2 approximately 8.4) is consistent with antagonism at the micro-opioid receptor in myenteric neurons. These results indicate that endomorphins directly activate the micro-opioid receptor in neurons, thus supporting the hypothesis that they are ligands mediating opioid actions in the nervous system. Endomorphin-induced micro-opioid receptor activation can be visualized by receptor endocytosis.

Analgesics, Opioid↗

PubMed1999

Tachykinin NK(1)receptor-mediated inhibitory responses in the guinea-pig small intestine.

We used in vivo, in vitro studies and immunohistochemistry to elucidate the mechanisms activated by tachykinin NK(1)receptors in evoking inhibitory motor response in the guinea-pig small intestine. In vivo, the selective NK(1)receptor agonist GR 73,632 produced a dose-dependent suppression of the distension-induced duodenal contractions, and a decrease of basal tone. These effects were reduced by pretreatment with the NK(1)receptor antagonist SR 140,333. In L-Nomega-nitro-L-arginine methylesther hydrochloride-pretreated animals, the suppressant effect of GR 73,632 on duodenal contractions was reduced, whereas the relaxation of the basal tone was unaffected. In vitro, GR 73,632 evoked a biphasic response consisting of a transient, tetrodotoxin-sensitive inhibitory effect followed by tetrodotoxin-resistant contractions. SR 140,333 blocked both inhibitory and excitatory motor responses induced by GR 73,632. NK(1)immunoreactivity was localized to myenteric and submucosal neurons and to interstitial cells of Cajal in the deep muscular plexus of the small intestine. NK(1)receptor-expressing neurons had Dogiel type I morphology and many of them were beta-nicotinamide adenine phosphate dinucleotide-diaphorase-positive, indicating they are inhibitory neurons. In conclusion, in the guinea-pig small intestine, NK(1)receptor stimulation evokes a myogenic excitatory motor response and a neurogenic inhibitory motor response that involves, at least in part, a nitrinergic pathway.

Adrenergic Agents↗

PubMed1998

Substance P (neurokinin-1) receptor is a marker of human mucosal but not peripheral mononuclear cells: molecular quantitation and localization.

Reciprocal communication between the immune system and the neuroendocrine system is mediated via a common chemical language of shared ligands and receptors. The neuropeptide substance P (SP) has been implicated as a mediator of immunomodulation. The evidence for substance P receptors on human lymphocytes is, however, controversial. The aims of the present study are to investigate substance P receptor (SPR) expression in human peripheral and mucosal mononuclear cells and to identify cellular sites of expression in human colonic mucosa. Using reverse-transcriptase PCR, we demonstrate that PBMC isolations are negative for SPR mRNA expression, whereas lamina propria mononuclear cell (LPMC) isolations express on average eight SPR mRNA transcripts per cell. In situ hybridization performed on surgically resected colonic tissue confirms the expression of SPR mRNA in LPMC in vivo. SPR mRNA signal was detected in LPMC, lymphoid follicles, and epithelium. The complementary technique of immunohistochemistry gave a similar distribution of SPR expression that colocalized with CD45 immunoreactivity. Dual-fluorochrome flow cytometry revealed SPR expression by CD4, CD45RO, CD45RA, CD8, CD19, and CD14 LPMC subsets, but not PBMC. Our findings suggest that SPR expression is distinctive of human colonic mucosal mononuclear cells and support a direct role for SP in mucosal immunomodulation.

Biomarkers↗

PubMed1998

Endomorphin-1 and endomorphin-2 activate mu-opioid receptors in myenteric neurons of the guinea-pig small intestine.

The novel opioid tetrapeptides, endomorphin-1 and endomorphin-2, recently isolated from bovine and human brain bind with high affinity and selectivity to central mu-opioid receptors. In the digestive tract, a comprehensive pharmacological analysis of the receptors involved in endomorphin action has not been reported. In this study, we analyzed the effects of endomorphin-1 and endomorphin-2 on longitudinal muscle-myenteric plexus preparations (LMMPs) from the guinea-pig ileum. Both peptides (30 pM - 1 microM) inhibited (-log EC50 values: 8.61 and 8.59, respectively) the amplitude of electrically-induced twitch contractions in a concentration-dependent fashion, up to its abolition. Conversely, in unstimulated LMMPs, they failed to affect contractions to applied acetylcholine (100 nM). In stimulated LMMPs, the highly selective mu-opioid receptor antagonist, D-Phe-Cys-Tyr-D-Trp-Orn-Thr-Pen-Thr-NH2 (CTOP), caused a concentration-dependent (30 nM-1 microM), parallel rightward shift of endomorphin-1 and endomorphin-2 inhibitory curves, without depression of their maximum. Following Schild analysis, calculated pA2 values were 7.81 and 7.85, respectively, with slopes not different from unity. Concentration-response curves to both peptides were not affected by 30 nM naltrindole (a selective delta-receptor antagonist) or 30 nM nor-binaltorphimine (a selective kappa-receptor antagonist). These results demonstrate that endomorphins selectively activate mu-opioid receptors located on excitatory myenteric plexus neurons, and that they act as full agonists.

Acetylcholine↗

PubMed1997

Neurokinin 1 receptor expression in the rat retina.

Tachykinin (TK) peptides influence neuronal activity in the inner retina of mammals. The aim of this investigation was to determine the cellular localization of the neurokinin 1 receptor (NK1), whose preferred ligand is the TK peptide substance P (SP), in the rat retina. These studies used a polyclonal antiserum directed to the C-terminus of rat NK1. The majority of NK1-immunoreactive (IR) cells were located in the proximal inner nuclear layer (INL), and very rarely they were found in the distal INL. Some small and large NK1-IR somata were present in the ganglion cell layer. NK1-IR processes were densely distributed across the inner plexiform layer (IPL) with a maximum density over lamina 2 of the IPL. Immunoreactive processes also crossed the INL and ramified in the outer plexiform layer where they formed a sparse meshwork. NK1-IR processes were rarely observed in the optic nerve fiber layer. Double-label immunofluorescence studies with different histochemical markers for bipolar cells indicated that NK1 immunoreactivity was not present in bipolar cells. Together, these observations indicate that NK1 immunoreactivity is predominantly expressed by amacrine, displaced amacrine, interplexiform, and some ganglion cells. Double-label immunofluorescence experiments were also performed to characterize NK1-containing amacrine cells. Sixty-one percent of the gamma-aminobutyric acid (GABA)-IR cells, 71% of the large tyrosine hydroxylase (TH)-IR cells, and 100% of the small TH-IR cells contained NK1 immunoreactivity. In addition, most (91%) of the NK1-IR cells had GABA immunoreactivity. In contrast, vasoactive intestinal polypeptide-, TK-, choline acetyltransferase-, and parvalbumin-IR amacrine tells did not express NK1 immunoreactivity. Overall, the present findings suggest that SP acts directly upon several cell populations, including GABA-containing amacrine cells and ganglion cells, to influence visual information processing in the inner retina.

Animals↗

PubMed1997

Somatostatin 2A receptor is expressed by enteric neurons, and by interstitial cells of Cajal and enterochromaffin-like cells of the gastrointestinal tract.

Somatostatin exerts multiple effects by activating distinct G protein-coupled receptors. Here we report the cellular sites of expression of the somatostatin subtype 2A (sst2A) receptor in the rat enteric nervous system by using a C-terminus-specific, affinity-purified antiserum and immunohistochemistry. Antibody specificity was confirmed by the cell surface staining of human embryonic kidney 293 cells expressing the sst2A receptor, the lack of staining of cells expressing the somatostatin subtype 2B receptor, and the abolition of staining by preincubating the antiserum with the C-terminus peptide used for immunization, SSt2A(361-369). The SSt2A receptor antibody recognized a broad 80 kDa band on Western blots of membranes prepared from cells transfected with sst2A receptor cDNA; following receptor membrane deglycosylation, the antibody detected an additional 40 kDa band. In the enteric nervous system, the sst2A antibody primarily stained neurons of the myenteric and submucosal plexuses, and abundant fibers distributed to the muscle, mucosa, and vasculature. Immunoreactive staining was also observed in non-neuronal cells, including presumed interstitial cells of Cajal of the intestine and enterochromaffin-like cells of the stomach. Fibers expressing sst2A receptor immunoreactivity were often in close proximity to D cells of the gastric and intestinal mucosa. Colocalization of somatostatin and sst2A receptor immunoreactivities was not observed in endocrine cells nor in enteric neurons. Double-label immunohistochemistry revealed colocalization of sst2A and vasoactive intestinal peptide immunoreactivities in enteric neurons. The multiple types of cells expressing the sst2A receptor, including enteric neurons and non-neuronal structures, in addition to the relationship between somatostatin and sst2A receptor elements, provide evidence that the sst2A receptor mediates somatostatin effects in the gastrointestinal tract via neuronal and paracrine pathways.

Amino Acid Sequence↗

PubMed1997

GAP-43 mRNA and calcitonin gene-related peptide mRNA expression in sensory neurons are increased following sympathectomy.

Sympathectomy has been shown to result in an increased density of fibers immunoreactive for sensory peptides in peripheral targets innervated by both sensory and sympathetic neurons, providing evidence for functional interactions between sympathetic and sensory systems. These findings provided the background for examining the hypothesis that axonal outgrowth is induced from sensory neurons following sympathectomy. We examined the expression of GAP-43 mRNA, a specific marker for axonal outgrowth, in cervical (C3, C7, C8) and thoracic (T1, T2) dorsal root ganglia (DRG) of the rat following bilateral removal of the superior cervical ganglion, to assess whether the described increases in peptidergic afferent fibers reflected axonal outgrowth. In situ hybridization was used with 35S labeled riboprobes complementary to GAP-43 mRNA, and to calcitonin gene-related peptide (CGRP) mRNA, a marker for a major subset of thin-fiber sensory neurons. The density of GAP-43 mRNA nearly doubled by 18 h following sympathectomy and reached a threefold increase by 3 days. By 45 days following surgery, the GAP-43 mRNA level was still nearly twice that of normal animals, CGRP immunoreactivity was also examined: the density of fibers in the iris and cornea of sympathectomized animals was considerably greater from two weeks to 45 days following surgery, than in sham-operated controls. Concomitantly, there was a slight but significant increase in CGRP mRNA expression in T1 and C3 DRG 14 days postsympathectomy. Quantitative computerized image analysis demonstrated that GAP 43 mRNA expression in sympathectomized animals was 1.5 times greater in medium-sized DRG neurons and almost fourfold greater in small DRG neurons than in control rats. These results indicate that sympathetic denervation elicits axonal outgrowth in the population of sensory neurons that give rise to the small unmyelinated and thinly myelinated axons of peripheral nerves.

Animals↗

PubMed1997

Organization of the peripheral nervous system: autonomic and sensory ganglia.

Major advances have been made in our understanding of autonomic and sensory transmission and function during the past two decades. These include (i) the establishment of the role of sympathetic and parasympathetic ganglia in relaying neuronal information from the central nervous system to effector organs, (ii) the recognition that enteric ganglia, the third component of the autonomic nervous system, contain independent integrative circuits that control complex local activities, (iii) the evidence for local effector functions of primary sensory nerves in addition to their role in sensory transmission, and (iv) the discovery of plasticity of both autonomic and sensory neurons during disease states and inflammation. A major contribution to these new concepts has been the recognition that in both autonomic and sensory ganglia a variety of transmitters coexist in single neurons. Co-transmission is a widespread phenomenon that enables autonomic and sensory neurons to exert fine and highly regulated control of various functions such as circulation, respiration, digestion, and immune response. This chapter will focus on the general principles and specific features of autonomic and sensory ganglia, with a particular emphasis on their general organization and neurochemical properties. Classical concepts and modern principles of classification of autonomic and sensory ganglia are discussed.

Animals↗

PubMed1996

Vesicular monoamine transporter 2 expression in enteric neurons and enterochromaffin-like cells of the rat.

The cellular localization of the vesicular monoamine transporter 2 (VMAT2) in the rat digestive tract was investigated with immunohistochemistry. VMAT2-immunoreactivity (IR) was localized to neurons and fibers of enteric and pancreatic ganglia, to processes supplying the gut wall, the pancreas and blood vessels, and to enterochromaffin-like (ECL) cells in the gastric corpus, which contained calbindin-IR. Few VMAT2-IR cells were also found in the gastric antrum, but they did not contain gastrin-IR. VMAT2-IR was expressed in extrinsic sympathetic fibers as demonstrated by the elimination of a portion of VMAT2-IR processes by sympathectomy. The VMAT2-IR pattern is consistent with the overall distribution of biogenic amine cell groups in the digestive tract. Our results provide further evidence that VMAT2 is the vesicular amine transporter responsible for accumulation of monoamines into secretory vesicles of monoaminergic neurons and ECL cells.

Animals↗

PubMed1996

Agonist-selective endocytosis of mu opioid receptor by neurons in vivo.

Opiate alkaloids are potent analgesics that exert multiple pharmacological effects in the nervous system by activating G protein-coupled receptors. Receptor internalization upon stimulation may be important for desensitization and resensitization, which affect cellular responsiveness to ligands. Here, we investigated the agonist-induced internalization of the mu opioid receptor (MOR) in vivo by using the guinea pig ileum as a model system and immunohistochemistry with an affinity-purified antibody to the C terminus of rat MOR. Antibody specificity was confirmed by the positive staining of human embryonic kidney 293 cells transfected with epitope-tagged MOR cDNA, by the lack of staining of cells transfected with the delta or kappa receptor cDNA, and by the abolition of staining when the MOR antibody was preadsorbed with the MOR peptide fragment. Abundant MOR immunoreactivity (MOR-IR) was localized to the cell body, dendrites, and axonal processes of myenteric neurons. Immunostaining was primarily confined to the plasma membrane of cell bodies and processes. Within 15 min of an intraperitoneal injection of the opiate agonist etorphine, intense MOR-IR was present in vesicle-like structures, which were identified as endosomes by confocal microscopy. At 30 min, MOR-IR was throughout the cytoplasm and in perinuclear vesicles. MOR-IR was still internalized at 120 min. Agonist-induced endocytosis was completely inhibited by the opiate antagonist naloxone. Interestingly, morphine, a high-affinity MOR agonist, did not cause detectable internalization, but it partially inhibited the etorphine-induced MOR endocytosis. These results demonstrate the occurrence of agonist-selective MOR endocytosis in neurons naturally expressing this receptor in vivo and suggest the existence of different mechanisms regulating cellular responsiveness to ligands.

Animals↗

PubMed1996

Cellular localization of Pan-trk immunoreactivity and trkC mRNA in the enteric nervous system.

The members of the trk family of tyrosine receptor kinases, trkA, trkB, and trkC, are the functional receptors for neurotrophins, a family of related neurotrophic factors. In this study, we investigated 1) the distribution of neurotrophin receptors in the developing and adult rat digestive tract with a pan-trk antibody that recognizes all known trks and 2) the cellular localization of trk-encoding mRNAs in the adult gut with single-stranded RNA probes specific for trkA, trkB, and trkC. In the developing myenteric plexus, trk immunoreactivity was present at embryonic day (ED) 14. Cells and fibers immunoreactive for trk could be visualized in the myenteric plexus at ED 16. At this age, dense staining was found in thick bundles of fibers in proximity to the myenteric plexus in the longitudinal muscle and in association with blood vessels in the mesentery. At ED 18, trk immunoreactivity was also seen in thin processes running from the myenteric plexus into the circular muscle, and in fibers and cells in intrapancreatic ganglia. By ED 20, immunoreactive staining was quite dense in both the myenteric and submucosal plexuses. At birth, virtually all enteric ganglia displayed strong trk immunoreactivity; the intensity of the staining at this age made it difficult to discern individual cells. During postnatal development, there was a decrease in cell body staining and an increase in the density of trk-containing fibers that became widely distributed to the gut wall and pancreas. The adult pattern of trk immunoreactivity was established between postnatal days 5 and 10. In adults, trk immunoreactivity was found in numerous enteric and intrapancreatic ganglion cells and in dense networks of fibers innervating all the layers of the gut, the pancreas, and vasculature. The trkC mRNA was expressed in adult enteric ganglion cells of both the myenteric and submucous plexus. By contrast, the trkA and trkB mRNAs could not be detected in enteric ganglia. All three trk mRNAs were expressed in dorsal root ganglia, which were used as positive controls. The density and wide distribution of trk immunoreactivity together with its persistence in adulthood support the concept that neurotrophins play a broad role in the digestive system from development through adult life, perhaps being involved in differentiation, phenotypic expression, and tissue maintenance. The presence of trkC mRNA in enteric neurons along with recent evidence that neurotrophin-3 plays a role in the development of the enteric nervous system suggest that trkC and neurotrophin-3 are a major neurotrophin system in the gastrointestinal tract.

Animals↗

PubMed1996

Calcitonin gene-related peptide innervation of the rat hepatobiliary system.

The digestive system is densely innervated by calcitonin gene-related peptide (CGRP)-immunoreactive neurons. The present study investigated a) the distribution and origin of CGRP-immunoreactive fibers in the rat hepatobiliary tract, and b) their relation with substance P/tachykinin (SP/TK) immunoreactivity using immunohistochemical and radioimmunoassay techniques. CGRP-containing fibers form dense networks in the fibromuscular layer of the biliary tree and surrounding the portal vein. Thin, varicose fibers are present at the base of the mucosa of the ducts. In the liver, labeled fibers are restricted to the portal areas and the stromal compartment. Neonatal treatment with capsaicin, a neurotoxin for primary afferent neurons, or celiac/superior mesenteric ganglionectomy depletes CGRP-containing fibers in the biliary tract, and reduces those associated with the portal vein. In contrast, subdiaphragmatic vagotomy does not appreciably modify the density of these fibers. Radioimmunoassay studies show a reduction of CGRP-immunoreactive contents in the biliary tract and portal vein by 84% and 65%, respectively, following capsaicin treatment, and by 80% and 66%, respectively, following ganglionectomy. By contrast, CGRP concentrations in vagotomized animals are comparable to those of controls. Most CGRP-positive fibers appear to contain SP/TK immunoreactivity, as indicated by double-label studies. These results demonstrate that the rat hepatobiliary tract is prominently innervated by CGRP- and CGRP/SP/TK-immunoreactive fibers, which are likely to originate from spinal afferent neurons. The abundance of these fibers and their association with a variety of targets are in line with the involvement of these peptidergic visceral afferents in regulating hepatobiliary activities, including hemodynamic functions of the hepatic vasculature.

Animals↗

PubMed1996

Monoclonal antibody to VIP: production, characterization, immunoneutralizing activity, and usefulness in cytochemical staining.

A very stable cell line has been generated that produces monoclonal antibody to VIP designated as CURE.V55. This hybridoma was produced by fusion of spleen cells from an immunized Robertsonian mouse containing the translocated 8.12 chromosome with FOX-NY myeloma cells that are APRT deficient. VIP monoclonal antibody producing cell line #55 was selected by limiting dilutions using thymocytes as feeder layers. Ascites fluid containing high concentration of VIP monoclonal antibody was produced from pristane-primed BALB/c mice. Ascites fluid contained approximately 20 mg/ml IgG and bound 50% of 2 fmol 125I-VIP at a final dilution of 1:50,000. Binding of this IgG1 antibody was inhibited by 50% at 5 nM concentration of either VIP 1-28 or VIP 7-28. Protein-A purified IgG of this antibody, used in a concentration of 30 mg/kg per rat (IV), completely reversed the inhibitory effect of gastric corpus contractions induced by intravenous injection of VIP (10 nmol/kg) in sodium pentothal-anesthetized rats. A control anti-keyhole limpet hemocyanin monoclonal antibody did not alter the stimulatory effect of VIP on gastric corpus contractions. Immunohistochemistry showed that this VIP monoclonal antibody stains neurons, and nerve fibers in human and rat gallbladder and the sphincter of Oddi as previously described with our polyclonal antiserum.

Animals↗

PubMed1996

Impairment of the gastric hyperemic response to luminal acid in cirrhotic rats.

Liver cirrhosis impairs gastric mucosal resistance to luminal acid in humans and in animal models. Because we have previously shown that pentagastrin enhances defensive as well as aggressive factors implicated in mucosal injury, we examined the hypothesis that the pentagastrin-mediated enhancement of mucosal defense mechanisms may be impaired in cirrhotic rats. Increased acid backdiffusion and susceptibility to gross mucosal injury, associated with an elimination of the hyperemic response to gastric barrier disruption, was observed in cirrhotic rats. In in vivo microscopic studies in anesthetized rats, cirrhosis had no effect on pentagastrin-associated enhancement of mucus gel thickness or baseline gastric mucosal blood flow, although baseline mucus gel thickness was decreased. Cirrhosis did, however, abolish the luminal acid-related hyperemic response to pentagastrin, which was associated with impaired intracellular pH homeostasis during acid superfusion. Cirrhosis did not alter submucosal calcitonin gene-related peptide immunoreactive nerves. We conclude that acid backdiffusion and pentagastrin-associated hyperemic responses are important mucosal defensive factors that are specifically impaired by cirrhosis.

Acids↗

PubMed1996

Action of sensory neurons in an experimental at colitis model of injury and repair.

The influence of sensory nerves on inflammation and healing was studied in a rat colitis model at different stages of inflammation. Studies were performed in animals with and without ablation of sensory nerves, which was achieved by pretreatment with the neurotoxin capsaicin. Colitis was induced by a rectal enema containing trinitrobenzenesulfonic acid (50 mg/kg) in 50% ethanol. Severity of inflammation was markedly increased 3 and 7 days after induction of colitis in the capsaicin-pretreated group compared with the vehicle group as determined by a macroscopic damage score (at 3 days, 12.0 +/- 0.7 vs. 7.5 +/- 1.5; at 7 days, 12.2 +/- 0.8 vs. 6.5 +/- 0.8; P < 0.05), by histology (ulceration score at 3 days, 82 +/- 12 vs. 40 +/- 11%; at 7 days, 92 +/- 5 vs. 46 +/- 13%; P < 0.05), and by myeloperoxidase activity (at 3 days, 133 +/- 30 vs. 42 +/- 14 U/mg protein; at 7 days, 76 +/- 11 vs. 39 +/- 11 U/mg protein; P < 0.05). There was no significant difference in the severity of colitis 14 and 21 days after induction of colitis between the capsaicin-pretreated group and the vehicle group. These data suggest that, in this model, sensory nerves have an important protective function in the acute and subacute phases of inflammation but do not seem to play a significant role in the later stages of chronic inflammation.

Animals↗

PubMed1995

Cellular sites of expression of the neurokinin-1 receptor in the rat gastrointestinal tract.

In the digestive system, substance P is an excitatory transmitter to muscle, a putative excitatory neuro-neuronal transmitter, a vasodilator, and a mediator in inflammatory processes. Many of the biological effects of substance P are mediated by a high-affinity interaction with the tachykinin receptor neurokinin-1. The aim of the present study was to identify the sites of expression of this receptor in the rat stomach and intestine by immunohistochemistry with a polyclonal antiserum raised to the intracellular C-terminal portion of the rat neurokinin-1 receptor. Neurokinin-1 receptor immunoreactivity is present in a large population of enteric neurons. The relative density of these neurons along the gut is colon > ileum >> stomach. In the intestine, stained neurons have a smooth cell body with processes that can be followed within and between plexuses, and make close approaches to other neuronal cells, but do not appear to project outside the plexuses, suggesting that they are interneurons. In the stomach, neurokinin-1 receptor-immunoreactive neurons are infrequent and have a poorly defined and irregular shape. Neurokinin-1 receptor immunoreactivity is also localized to numerous non-neuronal cells in the inner portion of the circular muscle layer of the small intestine, which have the appearance of small dark smooth muscle cells or interstitial cells of Cajal. These cells are postulated to form a "stretch-sensitive" system with the deep muscular plexus and thus constitute an important site of regulation of muscle activity. Double labeling immunofluorescence was used to simultaneously localize neurokinin-1 receptor and substance P/tachykinin immunoreactivities. These experiments demonstrate that in the enteric plexuses, substance P/tachykinin-immunoreactive varicose fibers encircle the cell bodies of most neurokinin-1 receptor-containing neurons, and in the inner portion of the circular muscle layer of the small intestine they lie close to neurokinin-1 receptor-immunoreactive non-neuronal cells. In addition, some enteric neurons express both neurokinin-1 receptor and substance P/tachykinin immunoreactivities. The present study provides strong evidence that the neurokinin-1 receptor is the tachykinin receptor mediating the actions of substance P on enteric neurons and smooth muscle.

Animals↗

PubMed1995

Developmentally regulated expression of alpha- and beta-calcitonin gene-related peptide mRNA and calcitonin gene-related peptide immunoreactivity in the rat inferior olive.

Immunohistochemical methods have revealed the transient neonatal expression of calcitonin gene-related peptide (CGRP) in olivocerebellar compartments, and it has been hypothesized that this peptide plays a role in the development of olivocerebellar connectivity. Furthermore, the distribution of the CGRP binding sites in the cerebellar cortex also favors this hypothesis. In this study, the pattern of postnatal expression of alpha- and beta-CGRP mRNAs in the inferior olive (IO) complex was analyzed using in situ hybridization histochemistry with RNA probes complementary to specific sequences of alpha- and beta-CGRP mRNAs, and the results were compared with the pattern of CGRP immunoreactivity. High levels of alpha-CGRP mRNA expression were found in specific subnuclei of the IO complex, i.e., the medial part of the dorsal fold of the dorsal accessory olive, the beta nucleus, the dorsal cap, the caudal third of the medial accessory olive, and the rostral part of the dorso-medial cell column; in the same subnuclei beta-CGRP mRNA was detected. The olivary distribution of the two CGRP mRNA coincided with that of CGRP immunoreactivity. The expressions of alpha-CGRP mRNA and CGRP immunoreactivity were restricted to the first 2 postnatal weeks, the peak being reached at the end of the first week; beta-CGRP mRNA was transiently expressed in the same olivary compartments, but only from postnatal day 6 to 9. In general, the alpha-CGRP signal was also more intense than the beta-CGRP signal. The present findings indicate that the alpha- and beta-CGRP mRNA expression in the olivary complex is under developmental control and restricted to specific olivocerebellar compartments. The data provide a basis for the transient expression of a CGRP olivocerebellar compartment and further support the hypothesis of a role for CGRP in the complex postnatal cerebellar phenomena of connectivity reshaping and synapse stabilization.

Animals↗